Intermolecular quenching of Edans/Dabcyl donor-acceptor FRET pair

Fluorescence resonance energy transfer (FRET) is one of the most sensitive techniques for monitoring biochemical events. The donor-acceptor pair 5-(2´-aminoethyl)aminonaphthalene sulfonic acid (Edans) and 4-[[4´-(N,N-dimethylamino)phenyl]diazenyl]benzoic acid (Dabcyl) has excellent spectral overlap between the fluorescence emission of the former and the absorption of the latter, resulting in efficient energy transfer. Strategies incorporating this donor-acceptor pair have been successfully applied to fluorescence-based assays of HIV-1 protease, human neutrophil elastase, human cytomegalovirus protease, and hepatitis C virus protease. The use of a FRET strategy is of particular importance considering our current interest in the design of a formulation able to respond to internal and external stimuli to locally release a cocktail of immunostimulating and chemotherapeutic drugs against colorectal cancer, using a fluorescence reporting system to monitor in real time the response to the treatment.

In this communication, considering a fluorescent probe of peptidic nature, the mechanism of quenching of the FRET pair Edans/Dabcyl was studied by performing photometric and fluorimetric measurements at pH = 7.5 in phosphate buffer. The quenching mechanism was studied using the Stern–Volmer plot, confirming that this FRET pair was involved in a dynamic quenching process.